Transforming growth factor-β induces loss of epithelial character and smooth muscle cell differentiation in epicardial cells

During embryogenesis, epicardial cells undergo epithelial-mesenchymal transformation (EMT), invade the myocardium, and differentiate into components of the coronary vasculature, including smooth muscle cells. We tested the hypothesis that transforming growth factor-β (TGFβ) stimulates EMT and smooth muscle differentiation of epicardial cells. In epicardial explants, TGFβ1 and TGFβ2 induce loss of epithelial morphology, cytokeratin, and membrane-associated Zonula Occludens-1 and increase the smooth muscle markers calponin and caldesmon. Inhibition of activin receptor-like kinase (ALK) 5 blocks these effects, whereas constitutively active (ca) ALK5 increases cell invasion by 42%. Overexpression of Smad 3 did not mimic the effects of caALK5. Inhibition of p160 rho kinase or p38 MAP kinase prevented the loss of epithelial morphology in response to TGFβ, whereas only inhibition of p160 rho kinase blocked TGFβ-stimulated caldesmon expression. These data demonstrate that TGFβ stimulates loss of epithelial character and smooth muscle differentiation in epicardial cells by means of a mechanism that requires ALK5 and p160 rho kinase.